Specific binding of a target in a sample may be accomplished using a variety of binders, for example antibodies, affibodies, anticalins, aptamers. High molecular weight binders such as antibodies are disfavored for some in vivo applications (e.g., for imaging applications) because they take a long time to migrate to the target site, demonstrate poor tissue penetration, remain in the subject for a long time, are primarily cleared through the liver and are more likely to produce an immune response than smaller binders. Accordingly, for some in vivo applications low molecular weight, fast-clearing binders are preferred for rapid migration to a target site, tissue penetration, and rapid clearance from the body.
One fast-clearing binder is the Affibody, a 7 KDa polypeptide based on the z-domain of protein A. Affibodies all share a common three-helix protein fold. Affibodies against novel targets are generated by randomizing the 13 amino acid residues in the IgG-binding surface using phage or yeast display techniques. Affinity-matured proteins are produced as either a single 7 KDa domain (monovalent Affibody) or as two tandem 7 KDa domains (bivalent Affibody).
Fast-clearing binders that are particularly well suited for some in vivo applications because of their tissue penetration and low antigenicity, may nonetheless clear a body in a way that interferes with desired applications. For example, clearance via the liver could interfere with liver-based imaging applications; clearance via the kidney of a radiolabeled binder might result in unacceptable radiation dose to the kidneys; or fast clearance from the blood may need to be slowed to achieve penetration of difficult-to-access tissues.
Accordingly, needs exist for methods to modify biokinetics of fast-clearing binders. Moreover, needs exist for fast-clearing biokinetically tuned binders for in vivo imaging that may be used for a variety of imaging modalities for preclinical or diagnostic imaging.